An integrated system consisting of imaging hardware, software, calibrants, and methodologies will be developed to visualize and quantitate the amount of Fluorescence Resonance Energy Transfer (FRET) occurring between donor and acceptor molecules in epifluorescence microscopy. Quantitative FRET imagery will supplant current qualitative imaging techniques as well as single-pixel 'spot' photometry techniques, such that the concentration of donor, concentration of acceptor, and the FRET efficiency can be determined for every pixel within a microscopic field of view. KAIROS' recently described 'FRET Beads' will enable the response function of the proposed FRET microscope to be determined in a straight forward, algorithmic manner. This recent innovation in the construction of FRET standards obviates the need for complex time- resolved methods and other methods, which require determining various radiometric responses within the microscope. In Phase 1, FRET Beads will be used to calibrate images containing spectrally distinct derivatives of Green Fluorescent Protein (GFP). In general, FRET microscopy will facilitate basic cell biology experiments involving tagged proteins (e.g., protein-protein interactions) as well as drug discovery (protein- small molecule interactions), where there is a need to quantitate the extent to which two molecules are interacting. Our commercial efforts will be directed towards developing turnkey instrumentation, software, and calibration standards to facilitate such discovery. PROPOSED COMMERCIAL APPLICATION: KAIROS will develop and market a Fluorescence Resonance Energy Transfer Microscope that is calibrated for a variety of FRET pairs useful in cell biology. The recent invention of 'FRET Beads' by KAIROS will enable this first-ever development of a steady-state FRET microscope. Commercialization will include development of the FRET microscope, imaging software, and a catalogue of calibrants for a variety of FRET pairs useful in cell biology and drug discovery.